Cervical cancer, caused by human papillomaviruses (HPVs), is a major public health problem, worldwide. About 230,000 women die of cervical cancer every year, the majority in developing countries. Although early detection via routine cytological screenings (Pap smears) and HPV testing have lowered both the incidence and mortality of cervical cancer, significant problems and barriers remain, including the low predictive value of current testing. As many as 3 million Pap smears are classified as inconclusive in the US every year leading to costly and invasive follow-up procedures and emotional stress in patients. MicroRNAs (miRNAs) are small, regulatory RNAs encoded by the plant, animal, and fungal genomes that act to inhibit expression of specific target genes. Recent studies have shown that miRNAs play key roles in many cellular processes, including development and differentiation. The role of miRNAs in the development of diseases and cancers has just begun to emerge. Furthermore, recent data show that many viruses encode miRNAs that regulate both viral and cellular gene expression to establish and maintain productive infection. Work at Ambion and in Golub's lab has shown that miRNAs are differentially expressed in specific types of cancers, providing the first evidence that miRNAs can be used to classify human tumors and develop diagnostic assays. Our hypothesis is that miRNAs are involved in the host-cell response to HPV infection and in HPV- induced cellular transformation, and that HPVs, themselves, encode miRNAs that are involved in these processes. We propose to investigate host cell and viral miRNAs involved in HPV infection for the purpose of better understanding the natural history of HPV infections and the early events that lead to the onset of cervical cancer. In Phase I, we will use a miRNA profiling system to analyze the host cell miRNA response to HPV infection and transformation in HPV-positive cell models and cervical biopsies. We will also explore the identification and validation of HPV-encoded miRNAs and determine their relevance to HPV infection and transformation. Phase II will encompass wider evaluation of the identified cellular and viral miRNAs in clinical samples as potential biomarkers and a diagnostic assay will be developed based on these miRNAs. [unreadable] [unreadable] [unreadable]